Induction of superoxide by 12‐O‐tetradecanoylphorbol‐13‐acetate and thapsigargin, a non‐phorbol‐ester—type tumor promoter, in peritoneal macrophages elicited from SENCAR and B6C3F1 mice: A permissive role for the arachidonic acid cascade in signal transduction

Abstract

Local production of reactive oxygen intermediates, e.g., superoxide anion, by tumor promoter-stimulated inflammatory macrophages (MPs) may contribute significantly to tumor development in classical models of two-stage chemical-induced carcinogenesis in murine skin. In the studies reported herein, peritoneal MPs elicited from phorbol-ester-sensitive SENCAR mice demonstrated a time- and dose-dependent release of superoxide anion (4-6 nmol/10(6) cells) when stimulated by 200 nM 12-O-tetradecanoylphorbol-13-acetate (TPA) in vitro; MP superoxide response was significantly inhibited (50-70%) by preincubation with 40 microM 1-(5-isoquinolinyl-sulfonyl)-2-methylpiperazine (H-7), a protein-kinase inhibitor. Alternatively, TPA-stimulated MPs derived from relatively resistant B6C3F1 mice generated negligible superoxide under the same conditions. A similar strain-dependent induction of superoxide was observed when MPs were stimulated with thapsigargin (TG), a tumor promoter previously shown to act independently of protein kinase C (PKC). TG-stimulated SENCAR MPs released a significant amount of superoxide (2-3 nmol/10(6) cells) that was not inhibited by H-7; MPs from B6C3F1 mice demonstrated negligible stimulation by TG. Preincubation of SENCAR MPs with 100 microM dibromoacetophenone, an inhibitor of phospholipase A2, completely suppressed the superoxide induced by TPA and TG stimulation. Like TPA, 50 microM 1-oleoyl-2-acetylglycerol, a diacylglycerol analogue and PKC activator, also induced a significant amount of superoxide from SENCAR MPs only. In parallel with the superoxide findings, TPA and TG stimulated significantly greater [3H]arachidonic acid release from prelabeled SENCAR MPs (a 32% and 48% increase, respectively, over unstimulated controls) relative to MPs from B6C3F1 mice. Two-dimensional gel-electrophoretic analysis indicated that TPA-induced phosphorylation of a 47-kDa protein (a presumed substrate for PKC previously linked to NADPH oxidase activation in guinea pig and human polymorphonuclear leukocytes) occurred in MPs from both SENCAR and B6C3F1 mice. Therefore, arachidonic acid production may be a common biochemical pathway by which phorbol-ester--and non-phorbol-ester--type tumor promoters activate MPs in SENCAR mice; such a response may be "permissive" for additive (or synergistic) interactions with PKC-driven signal pathways.

Transforming growth factor‐α expression in peritoneal macrophages elicited from SENCAR and B6C3F1 mice: Responses to lipopolysaccharide and 12‐O‐tetradecanoylphorbol‐13‐acetate

“…Therefore, strain-dependent differences in response were observed for TGF-a but, unlike the previously reported superoxide response [22], MPs derived from the relatively resistant B6C3F1 mice secreted greater amounts and only in response to LPS. Therefore, first and foremost, our results suggested that structural cells (e.g., epithelial cells and fibroblasts) may be the most significant contributors of the TGF-a detected in TPA-promoted SENCAR mouse skin [I 51.…”

“…Recent findings in our laboratory indicated that peritoneal MPs elicited from phorbol ester-sensitive SENCAR mice generated significant amounts of superoxide when stimulated by TPA in vitro; negligible responses were observed for MPs derived from relatively resistant B6C3F1 mice [22]. We hypothesized a similar strain-dependent secretion of TGF-a by TPA-stimulated MPs.…”

Inhibitory Effects of Curcumin and Tetrahydrocurcuminoids on the Tumor Promoter‐induced Reactive Oxygen Species Generation in Leukocytes in vitro and in vivo

“…Yoon et al advocated a close relationship between the generation of ROS, including O 2 − , by phagocytic cells in inflammatory processes and tumor promotion. 23) Kensler et al have hypothesized that the first treatment of mouse skin with TPA causes a chemotactic action, i.e., recruitment of neutrophils responsible for ROS generation induced by the second TPA treatment. 24) In fact, double application of TPA is required for excessive ROS production in mouse skin.…”

Ability of the non‐phorbol ester‐type tumor‐promoter thapsigargin to mimic the stimulatory effects of 12‐0‐tetradecanoylphorbol‐13‐acetate on ornithine decarboxylase activity, hydroperoxide production, and macromolecule synthesis in mouse epidermis In vivo

“…Whether or not they use PKC-dependent transductional pathways, Ca2+-mobilizing agents and TPA-or non-TPAtype tumor promoters may all increase the HPx-producing activity of the epidermis through AA release. After this report was submitted, another study indicated that the AA cascade may play a permissive role in the mechanisms by which TPA and TG stimulate the generation of 02by inflammatory macrophages elicited from promotion-sensitive SENCAR mice (Yoon et al, 1993). Although the Ca2+ ionophore and PL-A2 activator A23187 and the inhibitors of ,Ca2+ sequestration TG and CPA all mimic, at least partially, the HPx and DNA HPx production was determined 16 hr after the second of 2 TG or TPA treatments at a 48-hr interval.…”